A number of injectable protein-containing preparations are supplied to the market, in which various measures are taken to provide stabilized protein-containing preparations with low loss of active ingredients even after long-term storage. Protein-containing preparations are prepared by dissolving active ingredients and various additives such as diluents, solubilizers, excipients, soothing agents, buffering agents, sulfur-containing reducing agents, antioxidants, stabilizers, surfactants, etc. in a buffer.
A problem generally associated with the storage of proteins stored as concentrated solutions is their deterioration as exemplified by the formation of insoluble aggregates and that must be prevented.
For example, antibodies such as immunoglobulins, monoclonal antibodies and humanized antibodies are unstable proteins liable to physical or chemical changes such as association or aggregation under stresses of filtration, concentration and heating for removing viruses during the purification process.
A conventional method widely used for inhibiting deterioration of proteins to stably store them is stabilization by freeze-drying. However, it was necessary to add some agent for protecting against freezing to avoid denaturation or chemical changes that might be caused by mechanical stresses during freezing and freeze-drying.
A stabilization effect was found by adding as a stabilizer for inhibiting chemical or physical changes, polymers or polyols including proteins such as human serum albumin or purified gelatin or oligomers such as amino acids and surfactants. However, the addition of biopolymers such as proteins as stabilizers had problems such as the necessity of a very complex step for removing contaminants such as viruses derived from the stabilizers. Moreover, heat treatments for inactivating viruses sometimes caused problems such as association or aggregation due to heat stresses.
Interleukin-6 (IL-6) receptor is a ligand-binding protein having a molecular weight of about 80 KD to which IL-6 binds. Anti-IL-6 receptor antibodies were found to have a therapeutic effect on various IL-6-mediated diseases such as immune disorders, inflammatory diseases or lymphocyte tumors by blocking IL-6 signal transduction in immature myeloma cells to inhibit biological activities of IL-6 (Tsunenari, T. et al., Blood, 90:2437, 1997; Tsunenari T. et al., Anticancer Res. 16:2537, 1996). We also found that anti-IL-6 receptor antibodies have a therapeutic effect on immature myeloma cells (JP-A-8-099902).
We succeeded in the mass production of a reshaped humanized antibody hPM-1 as one of such anti-IL-6 receptor antibodies, and have tried to formulate this purified anti-IL-6 receptor antibody into pharmaceutical preparations.
Similarly to other protein preparations, an important factor for formulation of anti-IL-6R antibodies is chemical and physical stability. Especially, humanized anti-IL-6 receptor antibodies are unstable proteins liable to physical or chemical changes such as association or aggregation under stresses of filtration, concentration and heating for virus elimination during the purification process.
Thus, there is a demand for the development and commercialization of preparations containing an antibody, especially a humanized anti-IL-6 receptor antibody, which are stable even after long-term storage.
As for various physiologically active proteins, it is also necessary to establish preparation conditions and storage conditions for maintaining their structures and activities in order to supply them as pharmaceuticals in constant amount and with high quality. Especially, it would be desirable to develop a method for inhibiting formation of insoluble aggregates in protein solutions.